Introduction: NOD-like receptors (NLRs), of which NOD1 and NOD2 are the best characterised, are a class of intracellular pathogen recognition receptors involved in innate immune responses and are expressed in monocytic cells and the intestinal epithelium (1). Stimulation of NOD1 and NOD2 receptors with components of bacterial peptidoglycan results in cytokine production (2). Aberrant NLR expression has been associated with chronic inflammatory diseases (3). Mechanisms underlying NLR expression regulation are currently unknown. DNA methylation has been shown to modify gene expression and is effected by DNA methyltransferase enzymes (4). We hypothesised that DNA methylation may play a role in regulating NOD activity.

Method: To analyse the effect of DNA methylation on basal NOD1/2 expression; THP-1 monocytic or HCT116 intestinal epithelial cell lines were treated with a DNA methyltransferase 1 inhibitors; either 5μM 5-Azacytidine (5-Aza) or 500nM 5-Aza-2-deoxycytidine (5-Aza-dC) for 72 hours, after which NOD1/2 expression was analysed at the mRNA and protein level (n≥6) by quantitative polymerase chain reaction (QPCR) and western blotting, respectively. To assess NOD1/2 pro-inflammatory activity 5-Aza or 5-Aza-dC treated cells were subsequently stimulated for 6 hours with NOD1/2 ligands, after which expression of TNF-α and IL-6 were quantified at the mRNA level (n≥6) by QPCR. Data was analysed using independent t-test or two-way ANOVA analysis where appropriate, followed by appropriate post-hoc tests.

Conclusion: These findings suggest that NOD1 and NOD2 receptor expression and pro-inflammatory activity are epigenetically regulated and could potentially act as a novel drug target for chronic inflammatory disorders including inflammatory bowel diseases.

References:

1) Philpott, D.J., et al.,(2014). Nat Rev Immunol 14(1): 9-23.

2) Fenaux, P., (2005). Nat Clin Pract Oncol 2 Suppl 1: S36-44.

3) Chen, G., et al.,(2009). Annu Rev Pathol 4: 365-98.

4) Jones, P.A. (2012). Nat Rev Genet 13(7): 484-492.

Acknowledgements: This study was supported by funding from the Hardiman Scholarship.